title: Identification and conformer analysis of a novel redox-active motif, Pro-Ala-Ser-Cys-Cys-Ser, in Drosophila thioredoxin reductase by semiempirical molecular orbital calculation.
abstract: Mammalian thioredoxin reductases (TrxRs) contain selenium as selenocysteine (Sec) in the C-terminal redox center -Gly-Cys-Sec-Gly-OH to reduce Trx and other substrates; a Sec-to-Cys substitution in mammalian TrxR yields an almost inactive enzyme. The corresponding tetrapeptide sequence in Drosophila melanogaster TrxR (Dm-TrxR), -Ser-Cys-Cys-Ser-OH, endows the orthologous enzyme with a catalytic competence similar to mammalian selenoenzymes, but implementation of the Ser-containing tetrapeptide sequence SCCS into the mammalian enzyme does not restore the activity of the Sec-to-Cys mutant form (turnover number <2/min). MOPAC calculation suggested that the C-terminal hexapeptide Pro-Ala-Ser-Cys-Cys-Ser-OH functions as a redox center that alleviates the necessity for selenium in Dm-TrxR, and a mutant form of human lung TrxR that mimics this hexapeptide sequence showed improved catalytic turnover (17.4/min for DTNB and 13.2/min for E. coli trx) compared to the Sec-to-Cys mutant. MOPAC calculation also suggested that the dominant form of the Pro-containing hexapeptide is a C+ conformation, which perhaps has a catalytic advantage in facile reduction of the intramolecular disulfide bond between Cys497 and Cys498 by the N-terminal redox center in the neighboring subunit.
authors.affiliation: Department of Bioresources Chemistry, Graduate School of Natural Science and Technology, Okayama University, Okayama, Japan.
authors.forename: Mitsuhiko
authors.initials: M
authors.lastname: Kuwahara
authors.forename: Takashi
authors.initials: T
authors.lastname: Tamura
authors.forename: Kentaro
authors.initials: K
authors.lastname: Kawamura
authors.forename: Kenji
authors.initials: K
authors.lastname: Inagaki
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D055162:Biocatalysis; D003545:Cysteine / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000201:enzymology / Q000235:genetics*; D004926:Escherichia coli; D006801:Humans; D007700:Kinetics; D008168:Lung / Q000201:enzymology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012643:Selenium / Q000378:metabolism; D017279:Selenocysteine / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013880:Thioredoxin-Disulfide Reductase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D017279:Selenocysteine; D013880:Thioredoxin-Disulfide Reductase; D012643:Selenium; D003545:Cysteine